The roles played by metallothionein (MT) and more recently discovered lead-binding proteins (PbBP) from rat kidney and brain in regulating biological activity of Pb have been studied. MT mediates Pb inhibition of delta-aminolevulinic acid dehydratase (ALAD). Pretreatment of rats with Zn activated liver ALAD and attenuated the inhibition of ALAD by Pb in vitro and in vivo. MT isolated from Zn-pretreated rats injected with 203Pb disclosed that both Zn and 203Pb co-eluted in the MT fractions. Addition of purified liver ZnMT to purified bovine liver ALAD reaction mixtures increased enzyme activity by 2-fold and prevented inhibition of ALAD by Pb. Apothionein alone decreased the activity of Zn-activated ALAD and attenuated Pb inhibition of the enzyme. Studies of incubates containing 65ZnMT demonstrated that Zn was transferred from MT to ALAD. Fractionation of incubates containing 203Pb demonstrated that ZnMT sequestered Pb away from ALAD. These data suggest that MT may regulate the activity of some Zn-metalloenzymes by controlling Zn availability and that is may also alter the interaction of Pb with ALAD by decreasing the cytosolic pool of free Pb. Purification studies were conducted on the 10K PbBP from rat kidney. Amino acid analyses demonstrated a high glutamic and aspartic content with significant levels of phenylalanine and leucine but little cysteine. These data indicate that the 10K PbBP is not an MT but rather a novel carboxyl-rich, metal-binding protein with a unique amino acid composition. Western blot analyses using rabbit polyclonal antibodies to PbBP were conducted. The antibodies were specific for the PbBP and did not cross-react with MT or calmodulin and the protein was present in only kidney and urine. The data provide further evidence that PbBP is a novel renal protein which plays a major role in metal metabolism.